Optimizing skin disease diagnosis: harnessing online community data with contrastive learning and clustering techniques.

Skin diseases pose significant challenges in China. Internet health forums offer a platform for millions of users to discuss skin diseases and share images for early intervention, leaving large amount of valuable dermatology images. However, data quality and annotation challenges limit the potential of these resources for developing diagnostic models. In this study, we proposed a deep-learning model that utilized unannotated dermatology images from diverse online sources. We adopted a contrastive learning approach to learn general representations from unlabeled images and fine-tuned the model on coarsely annotated images from Internet forums. Our model classified 22 common skin diseases. To improve annotation quality, we used a clustering method with a small set of standardized validation images. We tested the model on images collected by 33 experienced dermatologists from 15 tertiary hospitals and achieved a 45.05% top-1 accuracy, outperforming the published baseline model by 3%. Accuracy increased with additional validation images, reaching 49.64% with 50 images per category. Our model also demonstrated transferability to new tasks, such as detecting monkeypox, with a 61.76% top-1 accuracy using only 50 additional images in the training process. We also tested our model on benchmark datasets to show the generalization ability. Our findings highlight the potential of unannotated images from online forums for future dermatology applications and demonstrate the effectiveness of our model for early diagnosis and potential outbreak mitigation.

Postamplifying Cas12a Activation through Hybridization Chain Reaction-Triggered Fluorescent Nanocluster Formation for Ultrasensitive Nucleic Acid Detection.

CRISPR/Cas12a-based biosensing is advancing rapidly; however, achieving sensitive and cost-effective reporting of Cas12a activation remains a challenge. In response, we have developed a label-free system capable of postamplifying Cas12a activation by integrating hybridization chain reaction (HCR) and DNA-copper nanoclusters (DNA-CuNCs). The trans-cleavage of Cas12a triggers a silenced HCR, leading to the in situ assembly of fluorescent DNA-CuNCs, allowing for the turn-on reporting of Cas12a activation. Without preamplification, this assay can detect DNA with a detection limit of 5 fM. Furthermore, when coupled with preamplification, the system achieves exceptional sensitivity, detecting the monkeypox virus (MPXV) plasmid at 1 copy in human serum. In a MPXV pseudovirus-based validation test, the obtained results are in agreement with those obtained by qPCR, reinforcing the robustness of this method. Our study represents the first effort to manipulate DNA-CuNC formation on HCR for highly sensitive and cost-effective reporting of Cas12a, resulting in an efficient synthetic biology-enabled sensing platform for biosafety applications.

Behavioral intentions of self-isolation and informing close contacts after developing mpox-related symptoms among young men who have sex with men in China.

The 2022 multi-country mpox outbreak raised public concern globally. Self-isolation and informing close contacts after developing mpox-related symptoms are critical measures in controlling the outbreak. This study investigated behavioral intentions of self-isolation and informing close contacts after developing mpox-related symptoms and associated factors among young men who have sex with men (YMSM) aged 18-29 years in China. The cross-sectional study was conducted among 2493 YMSM in six provincial regions in China from September 10th to 30th, 2022. Descriptive and logistic analyses were applied, using the intentions of self-isolation and informing close contacts after developing mpox-related symptoms as binary outcomes. The mean age of the participants was 24.6 (SD = 2.9) years. The prevalence of having intentions of self-isolation and informing close contacts after developing mpox-related symptoms was 88.6% (95% CI: 87.3%-89.9%) and 84.9% (95% CI: 83.5%-86.3%). Participants who were employed (adjusted odds ratio (AOR) = 1.474, 95% CI: 1.035-2.097; AOR = 1.371, 95% CI:1.002, 1.876), had higher mpox knowledge scores (AOR = 1.474, 95% CI: 1.035-2.097; AOR = 1.371, 95% CI: 1.002-1.876), and had higher perceived threats of mpox (AOR = 1.079, 95% CI: 1.030-1.130; AOR = 1.045, 95% CI: 1.002-1.090) were more likely to intend to self-isolate and inform close contacts. Participants who had MSM in-person gatherings in the past 6 months were more likely to intend to self-isolate (AOR = 1.392, 95% CI: 1.066-1.208). Participants with higher depression scores (AOR = 0.968, 95% CI: 0.948-0.989) and self-stigma (AOR = 0.975, 95% CI: 0.954-0.997) were less likely to intend to self-isolate and inform close contacts, respectively. Self-isolation and informing close contacts when developing disease-related symptoms are acceptable measures in response to mpox in China. Strengthening targeted risk communication and self-efficacy, raising disease knowledge, providing mental support, and reducing stigma toward the affected community are warranted.

Proctitis: An Approach to the Symptomatic Patient.

Proctitis is an inflammatory condition of the distal rectum that can be associated with common sexually transmitted infections (STIs), such as gonorrhea, chlamydia, and syphilis. For persons presenting with ulcerative findings on examination, in addition to syphilis, Mpox, lymphogranuloma venereum, and herpes simplex virus should be in the differential. Providers should also be aware that there are evolving data to support a role for Mycoplasma genitalium in proctitis. Performing a comprehensive history, clinical evaluation including anoscopy, and rectal nucleic amplification STI testing may be useful in identifying the cause of proctitis and targeting treatment.

A survey-based assessment of rates and covariates of mpox diagnosis and vaccination provides evidence to refine eligibility criteria for mpox vaccination among gay, bisexual and other men who have sex with men in the Netherlands.

Background: The 2022 multicountry mpox outbreaks predominantly affected gay, bisexual and other men who have sex with men (GBMSM) in non-endemic countries, including in the Netherlands. We conducted a survey-based assessment of the alignment between the risk factors associated with mpox diagnosis among GBMSM in the Netherlands and the eligibility criteria used in 2022 for vaccinating this group, with the aim to refine these criteria. Methods: An online self-report survey was conducted among adult GBMSM in the Netherlands between 29 July and 30 August 2022, corresponding to the first month of the Dutch mpox vaccination campaign. GBMSM were recruited via advertisements on social media and gay dating apps. Participants reported on their sexual behaviour, mpox diagnosis, and/or (initial) mpox vaccination since the start of the outbreak. Covariables of mpox diagnosis and vaccination were assessed using logistic regression analyses. Results: Of the 2,460 participants, 73 (3.0%, 95% CI 2.3-3.6%) were diagnosed with mpox and 485 (19.7%, 95% CI 18.1-21.3%) had received (initial) mpox vaccination. Using sample weighting, we estimated that, of the GBMSM population aged 18-80 years in the Netherlands, 1.1% (95% CI 0.7-1.6%) had been diagnosed with mpox and 7.8% (95% CI 6.8-8.9%) had received (initial) vaccination. HIV-PrEP use, living with HIV, reporting ≥20 sex partners in the past 12 months, and sex in sex venues/parties in the past 2 months were independent risk factors for mpox diagnosis. Except for sex in sex venues/parties, these variables were also independently associated with mpox vaccination. Conclusion: This study provides novel evidence regarding the degree to which the 2022 eligibility criteria for mpox vaccination align with the risk factors for mpox among GBMSM in the Netherlands. The findings contribute to a refinement of the eligibility criteria for mpox vaccination, to which sex in sex venues/parties should be added.

Integrated Network Analysis of Symptom Clusters Across Monkeypox Epidemics From 1970 to 2023: Systematic Review and Meta-Analysis.

BACKGROUND: The worldwide spread of monkeypox (mpox) has witnessed a significant increase, particularly in nonendemic countries. OBJECTIVE: We aimed to investigate the changing clinical symptoms associated with mpox from 1970 to 2023 and explore their interrelations. METHODS: In this systematic review and meta-analysis, 3 electronic databases were searched for English peer-reviewed studies conducted from January 1970 to April 2023 that reported any symptoms among confirmed mpox cases. We categorized the mpox epidemics into 3 periods: 1970-2002 (period 1, within the African region), 2003-2021(period 2, epidemics outside Africa), and 2022-2023 (period 3, worldwide outbreak). Following PRISMA guidelines, a meta-analysis was performed to estimate the pooled prevalence for each symptom. The correlation among symptoms was analyzed and visualized using network analysis. RESULTS: The meta-analysis included 61 studies that reported 21 symptoms in 720 patients from period 1, 39 symptoms in 1756 patients from period 2, and 37 symptoms in 12,277 patients from period 3. The most common symptom among patients from all 3 periods was rash (period 1: 92.6%, 95% CI 78.2%-100%; period 2: 100%, 95% CI 99.9%-100%; and period 3: 94.8%, 95% CI 90.9%-98.8%), followed by lymphadenopathy (period 1: 59.8%, 95% CI 50.3%-69.2%; period 2: 74.1%, 95% CI 64.2%-84.1%; and period 3: 61.1%, 95% CI 54.2%-68.1%). Fever (99%, 95% CI 97%-100%), enlarged lymph nodes (80.5%, 95% CI 75.4%-85.0%), and headache (69.1%, 95% CI 4%-100%) were the main symptoms in period 1, with a significant decrease in period 3: 37.9%, 31.2%, and 28.7%, respectively. Chills/rigors (73.3%, 95% CI 60.9%-85.7%), fatigue (68.2%, 95% CI 51.6%-84.8%), and dysphagia/swallowing difficulty (61.2%, 95% CI 10.5%-100%) emerged as primary new symptoms in period 2 and decreased significantly in period 3. Most other symptoms remained unchanged or decreased in period 3 compared to the former 2 periods. Nausea/vomiting had the highest degree of correlation (with 13 symptoms) and was highly positively correlated with lymphadenopathy (r=0.908) and conjunctivitis (r=0.900) in period 2. In contrast, rash and headache were 2 symptoms with the highest degree of correlation (with 21 and 21 symptoms, respectively) in period 3 and were highly positively correlated with fever (r=0.918 and 0.789, respectively). CONCLUSIONS: The manifestation of symptoms in patients with mpox has become more diverse, leading to an increase in their correlation. Although the prevalence of rash remains steady, other symptoms have decreased. It is necessary to surveil the evolving nature of mpox and the consequential changes in clinical characteristics. Epidemic countries may shift their focus on the potential association among symptoms and the high synergy risk. TRIAL REGISTRATION: PROSPERO Registration: CRD42023403282; http://tinyurl.com/yruuas5n.

Clinical Diagnosis Is Highly Predictive of Laboratory-Confirmed Mpox in a Sexual Health Clinic.

BACKGROUND: Promptly recognizing mpox can facilitate earlier diagnosis and appropriate treatment. How accurately clinicians can diagnose mpox based on clinical data and before receiving molecular test results is not known. METHODS: Leveraging public health and clinical data collected in Seattle-King County's Sexual Health Clinic (SHC) from July 29, 2022, to September 30, 2022, we analyzed the proportion of patients who received presumptive versus results-based tecovirimat when clinicians had a high, intermediate, or low suspicion for mpox after clinical evaluation. We calculated the sensitivity, specificity, and positive (PPV) and negative predictive value (NPV) of this approach against criterion standard mpox polymerase chain reaction (PCR) results. RESULTS: Of 321 patients evaluated for mpox in the SHC, median age was 34.5 years and 88% were cisgender men. Overall, 121 of 319 (38%) tested positive by mpox PCR. Clinicians had high suspicion for mpox in 122 patients and offered empiric tecovirimat to 92 (88%), of whom 85 (92%) tested PCR positive. Of 13 intermediate suspicion patients offered presumptive therapy, all accepted but none tested positive by PCR. The sensitivity, specificity, PPV, and NPV of high/intermediate clinical suspicion for mpox were 99%, 90%, 86%, and 99%, respectively. A higher proportion of people with HIV were diagnosed with mpox (57% vs. 36%, P = 0.01, χ2 test), and sensitivity and PPV of high/intermediate clinical suspicion in this subgroup were 100% and 86%, respectively. CONCLUSIONS: Clinical providers working in a high-volume, public SHC were able to both accurately identify and rule out mpox based on clinical examination before receiving PCR test results.

Amplification-free detection of Mpox virus DNA using Cas12a and multiple crRNAs.

Mpox virus (MPXV) is a zoonotic DNA virus that caused human Mpox, leading to the 2022 global outbreak. MPXV infections can cause a number of clinical syndromes, which increases public health threats. Therefore, it is necessary to develop an effective and reliable method for infection prevention and control of epidemic. Here, a Cas12a-based direct detection assay for MPXV DNA is established without the need for amplification. By targeting the envelope protein gene (B6R) of MPXV, four CRISPR RNAs (crRNAs) are designed. When MPXV DNA is introduced, every Cas12a/crRNA complex can target a different site of the same MPXV gene. Concomitantly, the trans-cleavage activity of Cas12a is triggered to cleave the DNA reporter probes, releasing a fluorescence signal. Due to the application of multiple crRNAs, the amount of active Cas12a increases. Thus, more DNA reporter probes are cleaved. As a consequence, the detection signals are accumulated, which improves the limit of detection (LOD) and the detection speed. The LOD of the multiple crRNA system can be improved to ~ 0.16 pM, which is a decrease of the LOD by approximately ~ 27 times compared with the individual crRNA reactions. Furthermore, using multiple crRNAs increases the specificity of the assay. Given the outstanding performance, this assay has great potential for Mpox diagnosis.

Identification of homologous protein models via 3D comparisons using predicted structures.

Recent advances in protein structure prediction enable 3D homology alignment and domain annotation using tertiary structures. Here, we present a protocol to identify homologous structures and annotate protein domains through in silico comparisons using the AlphaFold database. We describe steps for downloading and installing PyMOL software, preparing the query structure, and conducting a 3D homology search. The example provided highlights the application of this protocol in reevaluating an mpox viral protein annotation. For complete details on the use and execution of this protocol, please refer to Pan et al. (2023).1.

Prospective screening for monkeypox infection among users of HIV pre-exposure prophylaxis in Denmark.

INTRODUCTION: During the 2022 outbreak of mpox (previously called monkeypox), which primarily affected Gay, Bisexual, and other Men who have Sex with Men (GBMSM), testing was mainly limited to individuals with symptoms of infection. Although sporadic cases of mpox continue to be diagnosed in Denmark, the feasibility of screening asymptomatic high-risk populations, such as those using HIV pre-exposure prophylaxis (PrEP), is still unknown. METHODS: During the autumn of 2022, a rectal swab test for mpox PCR was included in the routine sexually transmitted infections (STI) screening for PrEP users. RESULTS: The screening included 224 asymptomatic men with a median age of 36.5 years. One patient (0.4%) tested positive for mpox. Ten (4.5%) and nine (4.0%) had chlamydia and gonorrhea, respectively. DISCUSSION: Our study demonstrates that screening for mpox is feasible in two Danish PrEP clinics.

Fabrication of label-free immunoprobe for monkeypox A29 detection using one-step electrodeposited molybdenum oxide-graphene quantum rods.

Monkeypox is a zoonotic viral infection caused by the monkeypox virus (MPXV), which belongs to the Poxviridae family of the Orthopoxvirus (OPXV) genus. Monkeypox is transmitted from animals to humans and humans to humans; therefore, the accurate and early detection of MPXV is crucial for reducing mortality. A novel graphene-based material, graphene quantum rods (GQRs) was synthesized and confirmed using high-resolution transmission electron microscopy (HR-TEM) and atomic force microscopy (AFM). In this study, molybdenum oxide was electrodeposited and one-pot electrodeposition of MoO3-GQRs composite on carbon fiber paper (CFP) enabled by an antibody (Ab A29)/MoO3-GQRs immunoprobe was developed for the early diagnosis of MPXV protein (A29P). Several studies were conducted to analyze the MoO3-GQRs composite, and the prepared Ab A29/MoO3-GQRs immunoprobe selectively bound to the A29P antigen that was measured using differential pulse voltammetry (DPV) analysis and impedance spectroscopy. The antigen-antibody interaction was analyzed using X-ray photoelectron spectroscopy. DPV analysis showed a wide linear range of detection from 0.5 nM to 1000 nM, a detection limit of 0.52 nM, and a sensitivity of 4.51 µA in PBS. The prepared immunoprobe was used to analyze A29P in serum samples without reducing electrode sensitivity. This system is promising for the clinical analysis of A29P antigen and offers several advantages, including cost-effectiveness, ease of use, accuracy, and high sensitivity.

Repeat testing of mpox specimens with late CTs improves detection of potential false positive cases.

The mpox pandemic necessitated the rapid development of clinical assays for monkeypox virus detection. While the majority of mpox specimens have high viral loads with corresponding early cycle threshold (CT) values, reports have indicated some specimens with late CT values can represent false positive results. To mitigate this risk, the Centers for Disease Control and Prevention (CDC) published an advisory recommending repeat testing of all specimens with CT values ≥34. However, limited experimental data were available to support this specific cutoff. In this study, we examine whether a more conservative approach in which all specimens with CT values ≥29 are repeated would improve the detection of potential false positive results. Compared to the CDC algorithm, our approach identified an additional 20% (5/25) of potential false positive results. To assess the impact of this cutoff on laboratory workload, we modeled the expected increase in test volume and turnaround time (TAT) relative to the CDC method. Using a lower repeat threshold, test volume increased by 0.7% while mean TAT increased by less than 15 minutes. Overall, a lower threshold than recommended by the CDC for repeating late CT mpox specimens may reduce the number of false positives reported while minimally impacting testing volume and TAT.

Detection of Mpox Virus in Seminal Fluids: Implications for Sexual Transmission.

The 2022 outbreak of the human mpox virus, formerly known as monkeypox, raised global health concerns with widespread transmission across multiple countries. Sexual transmission emerged as a significant mode of spread, particularly among high-risk groups like MSM and PLWH. This manuscript focuses on the implications of seminal fluids in the transmission of mpox. The virus has been detected in various bodily fluids, including semen, indicating the potential for sexual transmission. Studies have reported high positivity rates of mpox DNA in seminal fluids. Despite some concern about possible contamination due to genital lesions, the presence of replication-competent virus in seminal fluids has been confirmed and mpox virus was also detected in this specimen among people who engaged only in receptive sexual intercourse. Antiviral treatment with tecovirimat showed efficacy in reducing viral presence in semen with detection of the antiviral in this specimen. Virus clearance from semen is relatively rapid and parallels healing from infection, with no reported cases of seminal fluid relapses. The WHO recommendation to avoid condomless intercourse for 12 weeks after clinical healing still appears prudent. Continued research and surveillance are essential to understand viral dynamics and develop effective prevention measures to combat the spread of mpox through sexual transmission and protect key-populations.

Monkeypox virus-associated meningoencephalitis diagnosed by detection of intrathecal antibody production.

BACKGROUND: In the 2022 mpox-outbreak most patients presented with mild symptoms. Central nervous system (CNS) involvement has previously been described as a rare and severe complication of mpox; however, diagnostic findings in cerebrospinal fluid (CSF) analysis and neuroimaging studies have only been reported in one case previously. CASE PRESENTATION: We report a previously healthy 37-year-old man with mpox complicated by encephalitis. He first presented with painful skin lesions and genital ulcers; polymerase chain reaction (PCR) from the lesions was positive for mpox. Twelve days later he was admitted with fever and confusion. Neuroimaging and CSF analysis indicated encephalitis. The CSF was PCR-negative for monkeypox virus but intrathecal antibody production was detected. He spontaneously improved over a few days course and recovered fully. CONCLUSIONS: This case of mpox-associated encephalitis shows that CNS involvement in mpox infection may have a relatively mild clinical course, and that detection of intrathecal antibody production can be used to establish the diagnosis if CSF monkeypox virus-PCR is negative.

Review of virological methods for laboratory diagnosis and characterization of monkeypox virus (MPXV): lessons learned from the 2022 Mpox outbreak.

Monkeypox virus (MPXV), originally endemic in West Africa (Clade II) and Central Africa (Clade I), has recently emerged worldwide and has reinforced the need for rapid and accurate MPXV diagnostics. This review presents and critically discusses the range of virological methods for laboratory diagnosis and characterization of MPXV as well as related lessons learned and practical experience gained from the 2022 Mpox global outbreak. Real-time PCR is currently considered the diagnostic gold standard and ensures accurate and timely confirmation of suspected Mpox cases based on suspicious skin lesions, and digital PCR improves the precision of MPXV DNA quantification. Whole genome sequencing reveals the diversity within the Clade IIb outbreak and highlights the role of microevolution in the adaptation of the virus to the human host. Continuous genomic surveillance is important for better understanding of human-to-human transmission and prevention of the emergence of variola virus-like strains. Traditional virological methods such as electron microscopy and virus isolation remain essential for comprehensive virus characterization, particularly in the context of vaccine and antiviral drug development. Despite the current challenges, serological tests detecting a range of anti-MPXV antibodies are important adjunct diagnostic and research tools for confirmation of late-presenting or asymptomatic MPXV cases, contact tracing, epidemiological studies, seroepidemiological surveys, and better understanding of the role of IgG and neutralizing antibodies in the immune response to infection and vaccination. A multidisciplinary approach combining advanced molecular techniques with traditional virological methods is important for rapid and reliable diagnosis, surveillance, and control of the outbreak.

Identification of the myxobacterial secondary metabolites Aurachin A and Soraphinol A as promising inhibitors of thymidylate kinase of the Monkeypox virus.

Thymidylate kinase (TMPK) of monkeypox virus (MPXV) has emerged as a promising target for potential therapeutics due to its significant role in pyrimidine metabolism. While smallpox drugs are advised for treating monkeypox, the European Medicine Agency has sanctioned Tecovirimat due to its potent nanomolar activity. Nonetheless, there is a need for monkeypox-specific therapeutic options. In this work, we employed docking-based virtual screening and molecular dynamics (MD) simulations to identify myxobacterial secondary metabolites as promising anti-viral natural compounds capable of inhibiting thymidylate kinase. The computational pharmacokinetics and manual curation of top-scoring compounds identified six lead compounds that were compared in terms of protein-ligand contacts and protein-essential dynamics. The study shows that among the six candidates, Aurachin A and the Soraphinol analogues such as Soraphinol A and Soraphinol C remain very stable compared to other compounds, enabling the active site integrity via a stable dynamics pattern. We also show that other compounds such as Phenoxan, Phenylnannolone C, and 8E-Aurafuron B remain unstable and have a negative impact on the active site integrity and may not be suitable binders for TMPK protein. Analyzing the Aurachin A and Soraphinol A binding, the established hydrogen bonds with Arg93 and the conserved hydrophobic interaction with Tyr101 are consistent with previous experimental interactions. Additionally, a deeper insight into the indole and the aromatic ring interaction through π-π stacking and π-cation interactions, as well as the background of Aurachin A and Soraphinol A as a bioactive compound, has significant implications not only for its potential as a promising drug but also for directing future drug discovery efforts targeting the TMPK protein.

Monkeypox virus cytologic findings: An institutional experience with an emerging threat.

OBJECTIVES: Mpox is a viral disease caused by monkeypox, a highly contagious orthopoxvirus that resulted in a global outbreak beginning in spring 2022. Diagnosis is confirmed via polymerase chain reaction (PCR) testing of swabs from mucocutaneous lesions. Rare reports have documented the histologic changes of mpox lesions, but the cytologic features have not been described. We present the cytology findings of samples taken from swabs of mucocutaneous mpox lesions in 3 different patients. METHODS: The patients were all male, aged 55, 43, and 37 years, all with mpox confirmed by PCR testing. Swabs from chest (cases 1 and 2) and tongue (case 3) lesions were directly sampled and submitted in Aptima (case 1) or PreservCyt solution (cases 2 and 3). Liquid-based preps were prepared and stained using the Papanicolaou method. Specimens were assessed for viral cytopathic changes. RESULTS: All cases showed nuclear cytopathic changes (enlarged nuclei with open chromatin and prominent red nucleoli), 2 cases demonstrated multinucleated keratinocytes, and 1 case showed potential Guarnieri bodies. The chromatin margination and nuclear molding typical of herpesviruses was not appreciated. CONCLUSIONS: The cytopathic changes of monkeypox are not specific, but their recognition could prompt appropriate PCR testing. Monkeypox shows distinct cytologic changes compared with herpesviruses.

Burdening Perspectives and Treatment Modalities of Monkeypox: A Central Dogma.

The monkeypox virus (MPXV), belonging to the genus Orthopoxvirus, is responsible for causing the zoonotic illness known as Monkeypox. The virus was initially identified during an outbreak at a Danish Zoo in 1958 and has since been found to infect various mammal species worldwide. While African squirrels and other rodents are believed to be the primary hosts, determining the natural host has proven challenging. While MPXV can be studied using different animal models in laboratory settings, understanding its natural transmission routes remains complex and species-dependent. Recent developments have elevated the global health concern surrounding Monkeypox, leading to its designation as a Global Health Emergency of International Concern on 23 July 2022. Enhancing surveillance and case detection is crucial in navigating the unpredictable epidemiology of this re-emerging disease. Human infections with the monkeypox virus are becoming less frequent due to population growth and economic improvements. Monkeypox, similar to smallpox, can potentially be controlled and eradicated in the future through vaccines, appropriate treatment, and personal protective equipment.